Brain-derived neurotrophic factor is upregulated in rats with chronic pancreatitis and mediates pain behavior

Abstract

Objectives: We examined the role of brain-derived neurotrophic factor (BDNF) in the pathogenesis of pain in an experimental model of chronic pancreatitis (CP).

Methods: Pancreatitis was induced by retrograde infusion of trinitrobenzene sulfonic acid into the pancreatic duct of adult rats. Twenty-one days after injection, BDNF expression was examined in pancreas-specific dorsal root ganglia (DRGs) by immunohistochemistry, and protein levels were quantified from DRGs and spinal cord extracts. The effects of intrathecal infusion of a neutralizing antibody to BDNF on pancreatic hyperalgesia were assessed by the sensitivity of the abdominal wall to filament probing as well as the nocifensive behavior to electrical stimulation of the pancreas.

Results: Levels of BDNF in DRGs and spinal cords (T9-13) were significantly higher in trinitrobenzene sulfonic acid rats compared with controls, accompanied by an increase in the number of pancreas-specific neurons expressing BDNF immunoreactivity. Brain-derived neurotrophic factor antagonism suppressed phospho-tropomyosin-related kinase B receptor levels in the spinal cord and significantly reduced behavioral responses in rats with CP.

Conclusions: Brain-derived neurotrophic factor is upregulated in pancreas-specific primary afferent neurons in rats with CP, and BDNF antagonism is associated with a reduction of pain-related behavior in these animals, suggesting an important role for this neurotransmitter in the nociception of CP.

FIGURE 1

Chronic pancreatitis results in up-regulation of BDNF in sensory nerves. A, Cryosections of thoracic DRGs were fluorescently stained for BDNF presence. Representative picture of BDNF-positive neurons in a DRG section (top panel) and DiI-labeled pancreas-specific neurons (middle panel). The bottom panel shows a merged picture identifying both pancreas specific BDNF expressing neurons. Trinitrobenzene sulfonic acid–treated animals had a significantly higher percentage of BDNF-expressing pancreas-specific neurons compared to controls (50.9% vs 43.2%, P < 0.05). Scale bar, 50 μm. B, Levels of BDNF protein as measured by ELISA in thoracic T9-13 DRGs (top panel) and spinal cord (bottom panel) were also significantly higher in TNBS rats compared with controls (P < 0.006 vs P < 0.05, respectively).

FIGURE 2

Phosphorylated TrkB levels in spinal cord are upregulated in CP and normalize with intrathecal anti-BDNF. A, Phosphorylated TrkB levels quantified by Western blot and expressed as a ratio of β-tubulin expression normalized to rats without pancreatitis. B, Representative examples of the actual Western blots (normal= rats without CP).

FIGURE 3

Intrathecal neutralization of BDNF attenuates behavioral responses to noxious stimulation in CP. A, Von Frey filament response from before and after intrathecal infusion of control IgG in TNBS-treated rats. Plot of mean number of responses (of 10 applications per filament). These data show increase in responses after control IgG treatment (P < 0.001) in animals with CP. B, Von Frey filament response from before and after intrathecal infusion of anti-BDNF in TNBS-treated rats. Intrathecal infusion of anti-BDNF significantly reduced the response activity of animals with CP during VFF testing (P < 0.001). C, Response to graded electrical stimulation before and after intrathecal infusion of control IgG in TNBS-treated rats. Number of nocifensive responses during 5 minutes of electrical stimulation of pancreas before and after infusion of control IgG. Although a robust response was seen in electrical stimulation in all animals, no significant differences in response numbers were observed after infusion compared with experiments before infusion. D, Response to graded electrical stimulation from before and after intrathecal infusion of anti-BDNF in TNBS-treated rats. Number of nocifensive responses during 5 minutes of electrical stimulation of pancreas before and after infusion of anti-BDNF. Animals had significantly fewer responses to electrical stimulation after intrathecal infusion of anti-BDNF (P < 0.01).